44 - Endothelial cells (EC) regulate ROS generation in vascular smooth muscle cells (VSMC) from hypertensive rats

Abstract

ECs can regulate by reactive oxidant species (ROS), several functions of VSMC. This study aimed to verify the role of ECs on ROS generation and proliferation of VSMC from normotensive (2K), renal hypertensive rats (2K-1C) and SHR. We used transwell® technique in which ECs and VSMCs are co-cultured. We tested NO and ROS production by fluorimetric assay and VSMCs proliferation by the proliferating cell nuclear antigen (PCNA) expression in VSMC, extracted from 2K-1C, 2K rats and SHR in monoculture and co-culture with EC (HUVECs). NO production was lower in the VSMCs from 2K-1C (936.0 ± 11.0 FI, P˂0.05) than in 2K VSMCs (1,234.1 ± 20.0 FI) whereas ROS production was higher in 2K-1C VSMC (1,072.6 ± 17.0 FI, P˂0.05) than in 2K (958.02 ± 27.1 FI). In the same way, NO production was lower in the VSMCs from SHR (3,700.3 ± 12.0 FI, P˂0.05) than in VSMCs from normotensive (4,633.3± 22.0 FI). ROS production was higher in SHR VSMCs (4,460 ± 19.0 FI, P˂0.05) than in normotensive (3,317 ± 23.1 FI). ROS generation in co-culture of VSMCs and ECs for 72 h from 2K-1C aortas (938.0 ± 12.0 FI, P˂0.05) was lower than in monoculture of 2K-1C VSMCs (1,072.7 ± 17.3 FI) and SHR (1,312.0± 10.1,FI). NO levels were greater in VSMCs co-cultured with ECs (1,096.0 ± 29.0 FI, P˂0.05) than in monoculture (929.66 ± 13 FI). ROS production was higher in 2K-1C and SHR than in 2K VSMCs. ECs treated with NO-synthase L-NAME or ibuprofen (COX inhibitor), did not alter ROS formation in VSMC, but Angiotensin-II abolished ROS-induced effects of ECs. PCNA expression was similar in VSMCs in monoculture and in co-culture, whereas it was greater in VSMCs from 2K-1C and SHR in monoculture than in co-culture. Our data suggest that ECs protect VSMCs from hypertensive rats due to NADPH oxidase inhibition.